Neuropeptide Y-Y5 receptor

ABSTRACT

The invention provides isolated DNA molecules encoding the human, mouse and rat NPY-Y5 receptors. These isolated DNA molecules can be used to express the NPY-Y5 receptors in cells which can then be used to screen compounds for NPY agonist and antagonist activity.

The present invention relates to isolated DNA molecules which encode theneuropeptide Y-Y5 receptor. In addition the present invention relates tothe use of these molecules in the production of the neuropeptide Y-Y5receptor using recombinant technology and to methods of screening andtesting compounds for neuropeptide Y (NPY) agonist or antagonistactivity.

In developed affluent countries the prevalence of obesity is alarmingand it is now a massive contribution to morbidity and mortality inaddition to being socially disadvantageous. Fat deposition in theabdominal area is a particular problem in relation to risk of Type IIdiabetes and cardiovascular disease. However, until recently, themolecular mechanisms controlling appetite, energy expenditure andadiposity have been surprisingly ill-understood.

Obesity has well-known associations with non-insulin-dependent diabetes(NIDDM), hypertension, dyslipidaemia and coronary heart disease, as wellas less obvious links with diseases such as osteoarthritis and variousmalignancies; it also causes considerable problems through reducedmobility and decreased quality of life. Seven forms of rodent obesities,determined by single gene mutations, have been identified: yellow [Ay],adipose [Ad], diabetes [db], fat [fat], tubby [tub] and obese [ob] inthe mouse and fatty [fa] in the rat. The obese phenotypes caused bythese mutations differ in their age of onset, severity and the degree ofinsulin resistance. Similar phenotypes can also be seen in obese humans.Recently the molecular bases for some of these mutations has beenelucidated. Of these the [ob] gene product “leptin” has created the mostinterest. However, many other factors are also involved in regulatingenergy balance and body fat distribution. Four factors appear mostlikely to have an important role: these are neuropeptide Y (NPY),corticotropin releasing factor (CRF)/ACTH/glucocorticoids, insulin andgalanin. In particular, NPY and its receptors play an important role inthe regulation of appetite and in a related manner, obesity.

Neuropeptide Y (NPY) forms a family (called the pancreatic polypeptidefamily) together with pancreatic polypeptide (PP) and peptide YY(PYY),which all consist of 36 amino acids and possess a common tertiarystructure. Neuropeptide Y (NPY) receptors, members of the Gprotein-coupled receptor superfamily, are activated by one of the mostabundant peptides in the mammalian nervous system and subsequentlyinfluence a diverse range of important physiological parameters,including effects on psychomotor activity, central endocrine secretion,anxiety, reproduction, vasoactive effects on the cardiovascular systemand most importantly, potent effects on appetite. A number ofneuropeptides and classical neurotransmitters, including noradrenalineand serotonin, modulate ingestive behaviours. However, NPY stands outfrom the many neurotransmitters with experimental effects on food intakein being able to induce obesity. Injections of NPY into theparaventricular nucleus (PVN), have been shown to increase, in a dosedependent manner, feeding and drinking behaviour in the rat. A singleinjection of NPY can increase food intake several-fold for several hoursand is effective even during the light phase when rats usually eatlittle, and in animals that have already eaten to satiety. Consequently,NPY peptides are certainly among the most potent orexygenic substancesknown in either food deprived or satiated animals. Repeated NPYinjections into the PVN result in a massive and persistent feedingresponse and the rats ultimately develop obesity, with a true increasein body fat content. The importance of NPY as a mediator ofappetite/obesity regulation is further enhanced by the very recentreport that the obese gene product leptin inhibits NPY synthesis andrelease.

Injections of NPY into the paraventricular nucleus cause a prompt androbust increase in plasma ACTH levels and there is clear evidence thatNPY-induced ACTH secretion is mediated by corticotropin releasing factor(CRF). However, its mode of action as well as its interaction with CRFwithin the brain is largely unknown, as are its interrelationships withother hormones, such as insulin. Nevertheless an agent which increasesappetite and raises glucocorticoid levels might be important ingenerating central obesity.

Specific agonists and antagonists of NPY are therefore likely to be ofsubstantial benefit for therapy of a wide range of clinical disorders.As NPY possess a compact tertiary structure and different parts of themolecule are required for interaction with different subtypes of thereceptor, the logical developments of both agonists and antagonists iscritically dependent upon the availability and knowledge of specificreceptor structure.

It is presently known that NPY binds specifically to at least fivereceptors; Y1, Y2, Y3, Y4 and Y1-like (or “atypical Y1”). While it hasbeen demonstrated that NPY receptors couple to the adenylate cyclasesecond messenger system, it remains probable that additional NPYreceptor subtypes exist since there is evidence thatphosphatidylinositol turnover, cations, and arachidonic acid may alsofunction as second messengers for NPY.

Since NPY agonists and antagonists may have commercial value as, forexample, potential anti-hypertensive agents, cardiovascular drugs,neuronal growth factors, anti-psychotics, anti-obesity and anti-diabeticagents, the ability to produce NPY receptors by recombinant DNAtechnology would be advantageous. To this end, DNA molecules encodingY1, Y2, Y3 and Y4 have previously been isolated.

The present inventors have now isolated novel DNA molecules encoding thehuman, mouse and rat Y1-like (hereinafter referred to as NPY-Y5)receptors. Similar DNA molecules encoding human and rat NPY-Y5 have beendescribed in International (PCT) Patent Specification No. WO 96/16542,however, these encode receptors with, in the case of the human NPY-Y5,an additional 10 N-terminus amino acids, and, in the case of the rat 20NPY-Y5, an additional 11 N-terminus amino acids. Through analysis ofseveral cDNA clones and RT-PCR using specific primers for intron andexon sequences, the present inventors have confirmed that the human,mouse and rat NPY-Y5 receptor does not include these additional 10/11amino acids. The DNA molecules described in WO 96/16542 may thus exhibitlower expression rates over those of the present invention. In addition,the receptors encoded by the DNA molecules described in WO 96/16542, mayshow lower and possibly altered activity.

Thus, in a first aspect, the present invention provides an isolated DNAmolecule encoding an NPY-Y5 receptor having about 445 amino acids or afunctionally equivalent fragment thereof.

Preferably, the isolated DNA molecule encodes an human, mouse or ratNPY-Y5 receptor.

Most preferably, the isolated DNA molecule has a nucleotide sequencesubstantially corresponding or, at least, >80% (more preferably, >95%)homologous to that shown:

(i) at nucleotides 6291 to 7625 of FIG. 1 (SEQ ID NO:1),

(ii) at nucleotides 63 to 1397 of FIG. 2 (SEQ ID NO:3),

(iii) at nucleotides 115 to 1449 of FIG. 3 (SEQ ID NO:5), or

(iv) at nucleotides 73 to 1470 of FIG. 4 (SEQ ID NO:7).

The isolated DNA molecule may be incorporated into plasmids orexpression vectors, which may then be introduced into suitablebacterial, yeast and mammalian host cells. Such host cells may be usedto express the NPY-Y5 receptor encoded by the isolated DNA molecule.

Accordingly, in a second aspect, the present invention provides amammalian, yeast or bacterial host cell transformed with the DNAmolecule of the first aspect.

In a third aspect, the present invention provides a method of producingNPY-Y5 receptors comprising culturing the host cell of the second aspectunder conditions enabling the expression of the DNA molecule andoptionally recovering the NPY-Y5 receptor.

Preferably, the host cell is mammalian or bacterial. Where the cell ismammalian, it is presently preferred that it be a Chinese hamster ovary(CHO) cell, human embryonic kidney 293 cell or insect Sf9 cells.

In a preferred embodiment, the NPY-Y5 receptor is expressed onto thesurface of the host cell.

The DNA molecules of the present invention represent a NPY receptorwhich may be of interest both clinically and commercially as it isexpressed in many regions of the body and NPY affects a wide number ofsystems.

By using the nucleic acid molecules of the present invention it ispossible to obtain neuropeptide Y-Y5 receptor protein in a substantiallypure form.

Accordingly, in a fourth aspect, the present invention provides NPY-Y5receptor in a substantially pure form.

Preferably, the purified NPY-Y5 has an amino acid sequence substantiallycorresponding to any one of the amino acid sequences shown in FIG. 5.

In a fifth aspect, the present invention provides an antibody capable ofspecifically binding to an NPY-Y5 receptor.

In a sixth aspect, the present invention provides a non-human animaltransformed with a DNA molecule according to the first aspect of thepresent invention.

In a seventh aspect, the present invention provides a method fordetecting agonist or antagonist agents of NPY-Y5 receptor, comprisingcontacting a NPY-Y5 receptor or a cell transfected with and expressingthe DNA molecule of the first aspect with a test agent under conditionsenabling the activation of a NPY-Y5 receptor, and detecting an increaseor decrease in NPY-Y5 receptor activity.

In a further aspect, the present invention provides a nucleic acid probecomprising a nucleotide sequence of 10 or more nucleotides capable ofspecifically hybridising to a unique sequence within the DNA molecule ofthe first aspect.

In a still further aspect, the present invention provides an antisensenucleic acid molecule comprising a nucleotide sequence capable ofspecifically hybridising to an mRNA molecule which encodes NPY-Y5receptor so as to prevent translation of the mRNA molecule. Suchantisense nucleic acid molecules may include a ribozyme region tocatalytically inactivate mRNA to which it is hybridised.

The term “substantially corresponding” as used herein in relation to thenucleotide sequences shown in FIGS. 1 and 2 is intended to encompassminor variations in the nucleotide sequence which due to degeneracy inthe DNA code do not result in a change in the encoded protein. Further,this term is intended to encompass other minor variations in thesequence which may be required to enhance expression in a particularsystem but in which the variations do not result in a decrease inbiological activity of the encoded protein.

The term “substantially corresponding” as used herein in relation toamino acid sequences is intended to encompass minor variations in theamino acid sequences which do not result in a decrease in biologicalactivity the NPY-Y5 receptor. These variations may include conservativeamino Aid substitutions. The substitutions envisaged are:

G, A, V, I, L, M; D, E; N, Q; S, T; K, R, H; F, Y, W, H; and P,Nα-alkalamino acids.

The invention is hereinafter described by way of the followingnon-limiting example and further, with reference to the accompanyingfigures.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 provides the nucleotide sequence (SEQ ID NO:1) of a genomic DNAmolecule encoding the human NPY-Y5 receptor and includes the predictedamino acid sequence (SEQ ID NO:2).

FIG. 2 provides the nucleotide sequence (SEQ ID NO:3) of a EDNA encodingthe human NPY-Y5 receptor and includes the predicted amino acid sequence(SEQ ID NO:4).

FIG. 3 provides the nucleotide sequence (SEQ ID NO:5) of a cDNA encodingthe rat NPY-Y5 receptor and includes the predicted amino acid sequence(SEQ ID NO:6).

FIG. 4 provides the nucleotide sequence (SEQ ID NO:7) of a genomic DNAencoding the mouse NPY-Y5 receptor and includes the predicted amino acidsequence (SEQ ID NO:8).

FIG. 5 shows the degree of identity between the predicted amino acidsequence of the human (SEQ ID NOS:2 and 4), mouse (SEQ ID NO:6) and rat(SEQ ID NO:8) NPY-Y5 receptor proteins.

FIGS. 6a-f provide graphical results of binding assays conducted withCHO cells expressing NPY-Y5, Y5 ligands assayed were NPY, Leu 31 Pro 34NPY, PP, PYY, NPY 2-36 and PYY 13-36.

FIG. 7 provides graphical results of cAMP assays conducted on CHO cellsexpressing NPY-Y5 using the ligands NPY, Leu 31 Pro 34 NPY, PP, PYY andNPY 2-36.

EXAMPLE Experimental Procedures

cDNA and Genomic Library Screening

A human genomic P1 DNA library (Genome-Systems), a human foetal braincDNA library (P. Seeburg, University of Heidelberg) and a rathypothalamic cDNA library (Stratagene) were screened with a 632 bp³²P-labelled EcoRI/Pst1 fragment flanking exon 1C of the human NPY-Y1gene. Hybridisation with the probe was performed in a solutioncontaining 6×SSC, 5×Denhardt's solution, 0.1% SDS and 100mg/ml denaturedand sheared salmon sperm DNA at 60° C. for 16 h. Filters were washedtwice for 15 min in 2×SSC/0.1% SDS at 60° C. followed by a 15 min washin 0.1×SSC/0.1% SDS and exposed to X-ray film (Kodak, X-Omat) using anintensifying screen at −70° C. for 16h. P1 DNA from positive clones wasisolated according to the manufacturer's protocol. The DNA was digestedwith EcoRI, HindIII, BamHI and PstI then subcloned into the BluescriptSK vector (Stratagene) generating clones covering all of the human Y1and Y5 genes.

Nucleotide Sequence Determination

Supercoiled plasrnid DNA was alkaline-denatured and sequenced by thedideoxy chain termination method using T7 polymerase (Promega) (Sambrooket al., 1992). The oligonucleotide primers used initially werecomplementary to the flanking region of the vector and then based onsequences obtained in order to complete the sequence analysis.

Restriction Map Determination

P1 DNA was digested with restriction enzymes EcoRI, BamHI, HindIII,alone and in all possible combinations, electrophoresed on a 0.8%agarose gel, alkaline-denatured (0.4 M NaOH), capillary-transferredusing 0.4 M NaOH to Hybond N⁺ membranes and hybridised with severalspecific oligonucleotides, cDNAs and genoric DNA fragments obtained fromthe subcloning.

In Situ Hybridisation Analyses

Sense and antisense riboprobes to the human NPY-Y5 receptor weresynthesised using the DIG RNA Labelling Kit (SP6/T7) (BoehringerMannheim). cDNA corresponding to the coding region of the human NPY-Y5receptor was linearised and transcribed with either T7 (for antisenseriboprobe) or SP6 (for sense riboprobe) RNA polymerase according to themanufacturers instructions using digoxygenin labelled dUTP.

Postmortem brain tissue was obtained from a young adult male withoutneurological disease. Specific brain regions were dissected and fixed byimmersion in formalin for 36 hours and then embedded in paraffin. 6 mmserial sections were collected on slides subbed in chrom alum and storedat 4° C. until used. Sections were dewaxed in Histoclear (NationalDiagnostics) for 5 min, rehydrated in 100% , 70% and 50% alcohol for 2min each then washed in phosphate buffered saline (PBS) for 5 min.

Sections were pretreated for 10 min at room temperature with 5 mg/mlproteinase K (Boehringer Mannheim) in 5 mM Tris, pH 7.5, 5 mM EDTA.Sections were then washed twice with 0.1M glycine (in PBS) for 2 min,once in PBS then incubated for 1 h at room temperature in hybridisationbuffer: 2×SSPE, 50% formamide, 5% dextran sulfate, 1×Denhardt's reagent,100 mg/ml tRNA type X-SA (Sigma). Digoxigenin labelled riboprobes tosense and antisense DNA (500 ng) in 75ul of hybridisation buffer wereadded to the sections and hybridised at 42° C. for 18 h in a humidifiedenvironment using a Hybaid Omnislide PCR Thermal Cycler (IntegratedSciences). After hybridisation, sections were washed at room temperaturein 2×saline sodium citrate (SSC) buffer, 0.15M NaCl/0.015 M Na-citrate,pH 7.0 for 10 min, then 0.2×SSC for 30 min followed by treatment with 20mg/ml RNase [Sigma], in 10 mM Tris, pH 7.5, mM NaCl for 15 min at roomtemperature. After RNase treatment the slides were washed in 2×SSC for 5min at room temperature then 0.2×SSC at 37° C. for 30 min.

Tissues were processed for immunological detection by washing for 10 minin buffer A (100 mM Tris-HCl, pH 7.5, 150 mM NaCl), then incubated for30 min with a 2% blocking solution (Boehringer Mannheim) with 0.3%Triton X-100 in buffer A. The sections were then incubated for 2 hourswith an alkaline phosphatase-conjugated anti-digoxigenin antiserum(Boehringer Mannheim, diluted 1/500 in buffer A plus 0.5% blockingreagent), washed twice for 5 min each in buffer A followed by a wash in100 mM Tris-HCI, pH 9.5, 100 mM NaCl, 50 mM MgCl₂ for 2 min. Thelabelled probes were visualised using nitro blue tetrazolium andbromochloro-indoyl phosphate as substrates for 18 hours in the dark.Sections were washed for 10 min in 10 mM Tris-HCl, pH 8.0, 1 mM EDTA,then 3 quick washes in distilled water, mounted with Aquamount [Gurr]and examined using a Zeiss Axiophot microscope with Nomarsky opticsusing a blue filter.

Expression of NPY Y5

The rat Y5 receptor protein was expressed as follows: the mammalianexpression construct rpHz17 was made by subcloning a 1.9 kb fragmentcontaining the whole coding region and almost the entire 3′untranslatedregion of the rat NPY Y5 cDNA into the pPRC/CMV vector (Invitrogen). Theconstruct is under the control of the CMV promoter and contains theneomycin gene for selection. The expression construct rpHz17 wastransfected into mammalian cell lines CHO-K1 and HEK using a modifiedcalcium phosphate transfection method.

NPY-Y5 Binding Assay

The coding region of the NPY-Y5 receptor was subcloned in the pRC/CMVexpression vector and transfected into the chinese hamster ovary (CHO)K1 cell line by using a modified calcium phosphate transfection method.CHO cells were maintained under 5% CO₂ in Dulbecco's modified Eaglesmedium (DMEM)/Ham's F-12 medium (1:1) with 2 mM glutamine and 10% fetalcalf serum. Stably transfected cells were selected with neomycin andtested for the ability to bind NPY/PYY analogues. Transfected cells(1×10⁶) were incubated in 0.5 ml assay buffer [50 mM Tris-HCl, pH 7.4, 2mM CaCl₂, 5 mM KCl, 120 mM NaCl, 1 mM MgCl₂, 0.1% bovine serum albumin]in the presence of 0.05nM ¹²⁵I labeled NPY and increasing concentrationsof human NPY and related peptides. Cells were incubated for 3 hours at15° C. then layered onto 0.5 ml horse serum before being palleted in amicrocentrifuge for 4 min. Radioactivity was measured for 1 min in a γcounter. Results of binding assays involving CHO cells expressing NPY-Y5receptor are shown in Table 1, expressed as a percentage of the maximalspecifically bound radiolabeled NPY. Results are the pooled data fromthree separate binding curves with triplicate points.

TABLE 1 Peptide IC₅₀ (nM) Mean +/− SE NPY 7.2 +/− 0.2 Leu31 Pro34 NPY7.3 +/− 0.3 PP  21 +/− 4.3 PYY 25 +/− 4  NPY 2-36  27 +/− 3.4 PYY 13-36>1000

cAMP Assays

CHO cells expressing NPY-Y5 receptor were grown and maintained inDulbecco's modified Eagles medium: Hams F12 medium (1:1 v/v)supplemented with 2 mM L-glutamine and 10% (v/v) foetal calf serum at37° C. under an atmosphere of 10% CO₂ in humidified air in 150 cm³flasks. Experiments were performed in 24 well cluster dishes when cellshad reached confluence.

Inhibition of Forskolin-stimulated [³H]-cAMP Accumulation

Cell monolayers were incubated for 2 h at 37° C. in 1 ml/well of HEPESbuffered Hanks solution (HBH; 20 mM, pH 7.4) containing [³H]-adenine (74kBq/well). Prior to the addition of agonist, cells were incubated in 1ml/well HBH containing the phosphodiesterase inhibitor Ro 20-1724 for 30min. Agonists (in 10 μl HBH) were added to the assay system followingthe addition of forskolin (10 μM) and the incubation continued for 10min. The temperature of the incubation medium was maintained at 37° C.during these manipulations. Incubations were terminated by the additionof 50 μl conc. HCl to each well which lysed the cells. [³H]-cAMP contentof the supernatant buffer from each well was isolated by sequential ionexclusion Dowex-alumina chromatography. After the addition of emulsifierscintillator (15 ml), radioactivity was determined by liquidscintillation counting. Results are provided in Table 2.

TABLE 2 Peptide IC₅₀ Values (n = 3) NPY 163.7 ± 70.0 nM  PYY 45.1 ± 31.4nM PP 73.4 ± 47.4 nM [2-36]NPY 242.5 ± 171.4 nM Leu³¹Pro³⁴NPY 75.9 ±38.3 nM

Results

Identification of NPY-Y5 Receptor Gene

The cloning and characterisation of the 5′ upstream region of the humanNPY-Y1 receptor gene, while confirming the existence of severalalternative 5′ exons for the Y1 gene (Ball et al., 1995), also revealeda region of extensive homology with G-protein coupled receptors in exon1C, involving a partial open reading frame in the opposite orientation.Comparison of this 200 amino acid sequence, which contained parts of thethird intracellular loop and transmembrane domains VI and VII, with theGenbank database, identified the human NPY-Y1 receptor as the closestrelated receptor with 37% identity. Subcloning and sequencing of theentire 7 kb area between exon 1C and exon 1B of the Y1 gene confirmedthe presence of a gene encoding a novel NPY receptor subtype named Y5(FIG. 1). Screening of human fetal brain and rat hypothalamic cDNAlibraries with a 632 bp human genomic Y5 fragment under high stringencyidentified full length cDNA clones for both species. These sequencesencode a 445 amino acid long Y5 receptor (FIGS. 2 and 3). The humangenomic sequence (FIG. 1) shows two candidate initiator ATG codons,however analysis of several cDNA clones and RT-PCR using specificprimers for intron and exon sequences has established that one of theseATG codons (located 30 nucleotides upstream of the other ATG) is locatedwithin an intron. The overall identity between the human and rat NPY-Y5receptors after this correction is 89%. FIG. 5 shows that the degree ofidentity between the predicted amino acid sequence of the human and ratNPY-Y5 receptors.

The exon which encodes the 5′ untranslated region of the human Y5 geneis separated by a 2.7 kb intron from exon 2 and is located about 2.8 kbupstream of exon 1B of the NPY-Y1 gene. The close proximity of these two5′ exons orientated in opposite directions suggests a possibleco-regulation of transcription of both genes through a common promoterregion.

An interesting feature of the human Y5 gene, however, is the harbouringof exon iC of the NPY-Y1 gene within the coding region of the NPY-Y5gene. The 100 bp long exon 1C encodes, in its opposite strand, a part ofthe Y5 sequence containing most of the third intracellular loop of thereceptor protein. This cytoplasmic loop can vary significantly in sizebetween G-protein coupled receptors and is thought to be involved indetermination of the specificity of coupling to different G-proteincomplexes. In contrast to all other known NPY receptor subtypes, thisregion in the Y5 receptor is unusually large, consisting of about 150amino acids. In the corresponding region of the NPY-Y1 gene, shortlyafter the fifth transmembrane domain, a small 97 bp intron containing anin frame stop codon interrupts the coding region (FIG. 1) suggestingthat this noncoding region has gained two additional functions afterduplication. One is to encode part of the Y5 protein sequence and theother is to fulfil a regulatory function in tissue specifictranscription, as an alternatively spliced 5′ exon of the Y1 gene.Transcription activation of exon 1C certainly will have an effect on Y5expression, most likely inhibiting mRNA production. However, such amechanism may represent only one aspect of a regulatory interactionbetween these two receptor genes. The close proximity of exon 1B of theY1 gene and exon 1 of the Y5 gene suggests an additional controlmechanism(s) for the specific transcriptional activation of one or theother gene.

Pharmacological Characterisation of the Y5 Receptor

NPY binding analysis of CHO cell lines stably expressing the rat Y5receptor subtype show a ligand specificity and rank order of potency(NPY=NPY>PYY[Leu³¹,Pro³⁴]=NPY[2-36]=PP>>PYY[13-36]) indicative of a NPYreceptor with a Y1-like pharmacology, as well as responding strongly tothe feeding specific ligand NPY[2-36] (FIGS. 6a-f). The same profile ofselectivity for these different NPY analogues can be seen in the resultsobtained from experiments measuring the inhibition of adenylate cyclaseactivity (FIG. 7).

In situ Hybridisation Analysis

A comprehensive study was made of the distribution of the Y5 receptormRNA in hypothalamic regions of the human hypothalamus. Hybridisationwith a sense probe to Y5 showed no specific labelling, however,antisense probe showed extremely high expression of Y5 receptor mRNA isfound in large neurons of the paraventricular nucleus. High levels arealso found in the dorsomedial nucleus, supraoptic nucleus and in themamillary body as well as in the midline thalamic nuclei. Within anucleus the distribution was not always homogenous. For example in thedorsomedial region, clearly unlabelled large pyramidal neurons werefound mingled with labelled neurons, suggesting functionalspecialisation. Preliminary results for the Y1 receptor suggest that thehuman NPY-Y1 receptor has a similar distribution to that of the Y5receptor, however, with some identifiable differences supporting thetheory of a co-regulatory transcription activation of the two genes.

Expression of NPY-Y5

The expressed Y5 receptor protein appears to have a unique distributionand relative affinities for different NPY/PYY/PP analogues. It is alsoexpected that the Y5 receptor will be functionally unique, relative toother NPY receptors, and may be very important in, for example, thedevelopment of drugs for a number of conditions such as appetite/obesitydisorders, hypertension, locomotor problems, memory loss, sleepingdisorders, migraine and gastrointestinal (GI) and cardiovasculardisorders.

It will be appreciated by persons skilled in the art that numerousvariations and/or modifications may be made to the invention as shown inthe specific embodiments without departing from the spirit or scope ofthe invention as broadly described. The present embodiments are,therefore, to be considered in all respects as illustrative and notrestrictive.

REFERENCES

1. Ball, H. J., Shine, J. & Herzog, H. (1995). Multiple promotersregulate tissue-specific expression of the human NPY-Y1 receptor gene.J. Biol. Chem. 270, 27272-27276.

2. Sambrook, J., Fritsch, E. F. & Maniatis, T. (1992). Molecule cloning(A Laboratory Manual) 2nd ed. (Cold Spring Harbor Laboratory Press, ColdSpring Harbor, N.Y.).

8 1 8371 DNA Homo sapiens 1 ctgcagcggc cggggcgccc cgaggtacgg gctcccgcccctccctgcca acccctttcg 60 cgccgggtag gcctgcaccg aggggccgtg gcgggtccccgcgcgggctg cgagtctgcg 120 caggtccctg ggagcccgca cccgtctctg gtgccagggcgttgtcgggg gtcccaagag 180 agcggggtgg ggaaggtgaa gggagcgcgg ctggaaaaatggggattagg gtggcggaac 240 aggcacttgt caggagtgaa gagacagcgg agagggtactgggctgaatt ctttcgtgcc 300 gagcaggtcc ctccggttcc caactcaccc gggtggagcaggcgcgggcc gaacccggga 360 ggagagtgtc ggggatccgc gaaggagcct cctggggatggggcggggga tggacaaagc 420 gctgcccccg gctggacacg ctctggcgct agcccggctggcatccggag ctgggaacag 480 caccccgcgg ggtgcccggg tcagggctca acctagcgggtctctggcga ggccgggggc 540 gcagcccgcg gggcgccact caggccgtcc agctgccgcgcggtccagcg ctgacccgag 600 cccgggaggc agctgcgctc taaggtttgc gctcctgtttgcgaggtgtc ttcatataac 660 aaatgcgagc aataacaaac atccatagaa ctcgaattccagaaacggga attctttttt 720 ccaagttcac agacctttag ttaatctttt aaaggaactgaggcgttgtg ttggaccaaa 780 gccaaaacga ttttacctta caccatggaa aatagcctaaggctcttttc agcagaattt 840 ttggcagtcc gaatgcaatt tttagatttc agatttctcaagggaagaga aactctgctg 900 ttagaatttg gaagggaggg tggtgcatgc ctgtgtgtttgtcagctgag cagagctgta 960 tttatctttc caattcaaat tgtgccagat tctggctttaagaaaaaacc atgggaatat 1020 ttgagaacat ggaatcatgc tgctgttcca cgatcacagcaaaacagaca atagttgata 1080 ttgtatcatt gcaggaggaa aaagaattac atatattttattcttttgtg tgattgtcat 1140 cctttgtgaa aagaatgatg tgtattttca taaagcaaaaaattattcaa acaaagaaac 1200 cttatttaaa tgtacaagtc agacttttaa tatcctttgaattccctgca gttcctccta 1260 ttattcttga gaactatcta cttggttaaa atacttaaatctattcagaa ggtttcattt 1320 gtctaggtct cagatataga agagtttata agaaaattccagtaaacctt taaaaagata 1380 ttatttttta taagttgcca tagtttaata aagaacttttatttttcaca ctttttactc 1440 agagattaaa gttctgtgtt tcagcctgga aattctgatggtgggagata caactaatac 1500 aaaagagaat gagtaaatat agtaattagg tatgacaaaagtctcatgct gtcaatatca 1560 gatttcttgt caaataatat tccatgttaa aatattttttctctggctat atttcataat 1620 ttatatagca atttcagaag attcacatat atcattacttttataataga taaaatatgt 1680 tgcataaaaa tgacagcact cgtaataaca cttgttgaaatttggatttc cattgtaggt 1740 ctgctcattg tgttttcagg aaaaaggaag ggaaagggtaagtttaatgg aaaaaatcct 1800 gcttttttgt ttgttttttc atttaagtgc gttcctgtaccttgagtttt caagttaaat 1860 cttattgtac aaaattttcc taatgtttaa actaggccctggctaccagg aggcactttt 1920 aaaaaaacta cacgtccacc accacccctc ccccacccgccctccctgcc tccagcattt 1980 gcaatattca ttatttagtt gtaagaagaa attcttccttcattggagca aagattcaca 2040 gaatgttcat tctgtgcaga ctatatatta gatattacatgtgtgtatgt ttatgtggta 2100 gatggtgtgg ggtggggcta gagggagagc aggagaaagttgactacagt cacaccaaaa 2160 taaaatgaat aaatgagtgt tgaatgaatc aagtgctaagagagaatttt taaattgctt 2220 accaatctat cagtagctac ataagtattc attatattcagcagtaatgc atgtgtccat 2280 gctatagaga aataatatat tactatcagt caggagaatgccattcattt attaattcat 2340 tcatcatcca atttgggcct ttttatatct cagcaatctacagttactca gggtgtagag 2400 cttgaattaa tctatataga atattcttgg catagcaccttgcattagtc gtctttatgc 2460 ttagagcaga gcagagcacc tagcagaata tatgttcaataaatactttt tgaatgaata 2520 aaagaaggaa caactaatca ttcttagctg ttcattaatagaaggtgcct acccctttaa 2580 aattatatat aaattatctc tttcttaaaa tactcaaatgttttaaggaa tgaaagaagc 2640 atcctcagtt ttttctccag tgtccaatga atactcaagatggcatttat ttcatcttct 2700 tactaaggag atgtggtttt acaatttaat gcattcaatattttatgtgc atatatttaa 2760 aataaaagtt ttaataacag actgcacagt cgcggaaatggatatacttc ttttttcatt 2820 tacatttttt aaatgttgta aatatatctt acagttttagttgcatgttg cttgtgtgat 2880 agcctttatc aatgaagtta tccaaattta aagtgctaaactatctttat tgtctgtcta 2940 ggtatctcct cctcattgca ttttggggcc atttgaaacatctataattt caatggttct 3000 ctataaatgt atatataaag atacatatac acacatatatatgtacacac aaaaatatag 3060 tcatactcta tcctgaattt tcccacattg ccagaatgattcatttctgt tattttaaag 3120 caagggaaat taaactgctt ttctaaaacg attggtaagaaatatttact tagcatccac 3180 tatgtgtaat atgctttatt aaacatcatt tctagaatgaaaataattaa gagttttatc 3240 tccattcgaa tataatagag aggtctaacc acatggaatggagaaaaatc tgaattttag 3300 actcaaaact acattgtttc tattaccaca aattgtgctgcatcttctct ttcttcaaaa 3360 aattttggac agcaatttta cactaagtaa gtatcatccacagttacatg ttccaaaaag 3420 gcacaaagcc gttgtagaag gggccatcta atttctctcttgttcttgct taggtgttac 3480 aaggaaaggc tatcggtaac aactgacctg ccacaaagttagaagaaagg attgattcaa 3540 gaaagtaagt caagagaaga acaactaagc aggattgcagttacaagcag cctgtacaca 3600 attataaata taaataggat catgaataag ctgaattgagccaggggatc atcagaactc 3660 aggaaattag gcaaaagcac cagtcaaagc tgttttgattagaagcttgc tgacctatcc 3720 agagtaggtg ctgagaggcc attgactggg aatatgatgaataatatgat tcagtaggtc 3780 atgcgagtca cttttgtacc aggtgttctt tgtcattgaggcaatatcaa tgtaaattgt 3840 tggctagggt ctaagaatga atgaatacaa tcctaagtctttgaattaac ttatccttta 3900 aaaggatgta gttagcttcc agaaaataat ttggtcaacatagaatcact tgtagaagtt 3960 gtgaaaaact tgtaactttt ctcatagcac aatgatgactctgtcatcct gtttgaaact 4020 tgctacacat agaactgaag ttaaacttat ttgtaatgaatgtatgtaca caatagtatt 4080 tgccatttgg aaatttattg aacgaagacc tgcaggtccctcataaatta aagataacag 4140 tgtttactat taatttaaat aaacatgtat ttttatagttttagtataat tattcaatta 4200 tagatctaga aataagtaga taaacatata ttgataggtaacaaaagtgg ttttttaact 4260 atatatatca caatctctac gacaatgtat ttattggaattaatttcttt gttggtttgt 4320 gttttctgta ggaaattctt gttaaaaaaa cattaaagtggctgggcaca gtggttcatg 4380 cctctcatgc ctataatccc aacagtttgg gaggccaaggtgggaggttt acttgaggcc 4440 aggagtttga gaccagcctg ggcaacatag ccagaccccatctccacaaa aaatagaaag 4500 attagccaga tgtagtggca cgtgcctgta gtccacgtgcctgtagtcca gctgcttggg 4560 aggctgagat gagaggattg cttgagtcca ggcgttcaaggttacaatga gctgtggtca 4620 cactactgca ccccagcctg ggcaacagaa tgagaccctttttctaagaa aaataaaaag 4680 gtaaaaaaaa aaaaaagtcc tttttttttt aaacgagaggagggagtcct tttgcctctt 4740 attggtatgt tataggcaat ttagtgcttc atcaggcagtaggatcaaaa gtctaatatg 4800 tagaggtaaa tacgtaatgc cattgatgta tgacattaatttaatttgaa atgaagaaaa 4860 cttattaccg ggagttatat taatatcact gctacatttacgtttaaggt ataatgtttt 4920 ccttgaacaa tgaattcatt gactcgttca taagccaaaatctatacaca gtttttaaat 4980 taatcaacag gtgaaatttg attgtttgtt tttttaaaacgccaacagcc tgctagtctg 5040 tcagtggttg tcctaatcag agataatctg gcacatctcaaaccattgag gattggtcac 5100 agaaagatgt catcatccag cattgcgtcc acacagtcaacagtagagtt tgataaatat 5160 atttaatgag tgcctactat atgcatctgg gtcatgagatagtgatccta ttctcaagga 5220 gcataaattt gaacattgta cgaactaggt gatatttgttactagagttt tgtttgaacg 5280 ttttattctc tcataaacat ttatttaata cctgcagtgatgaagttact ctgccatgta 5340 ttgggatgga ttccaaagtg agtaagagat agtttctgcttttccattgc ttgtaaataa 5400 acaaggtaga tgggtaggca ttataatgca atgaaagcagattatgatat gtagcatcag 5460 acaactgtaa acagaatgta acaggagttc tgaagaggagatcatgtcca gccgagttga 5520 ccaggacaag tgacttttaa gtttggccta gattgagatagaaataaatg gaatttttat 5580 gataagatta tgtgactata ctacatacca ggtatattgacttggagaat aatattaatg 5640 agtgattgca aagcatgtat cttgaagttc ttgtctacatttgccttttt ctttccttac 5700 gttatttact acagaaattt taaaaatgca atctactaccttaacataaa ttaatacatc 5760 ttagaagtaa tgataaaatt aaatttacta taatcattattggctgatac ttgaattgcc 5820 cttggaacga gttaaaggta tcataaactt tctgggctgggcacggtgct cacgcctgta 5880 atcccagcac tttgggaggc cgaggcgggc ggatcacgaggtcaggagat cgagaccacg 5940 gtgaaacccg gtctctacta aaaatacaaa aaattagctgggcgcagtgg cgggcgcctg 6000 tagtcccagc tactcgggag gctgaggcag gagaatggcgtgaacccggg aggcggagct 6060 tgcagtgagc cgagatggcg ccacagcact ccagcctgggcgacagagcg agactccgtc 6120 tcaaaaaaaa aaaaaaaaaa aagatatcat aaacttccttaggagattaa taaggtcacg 6180 ggagctgatt gtaatattta gtttccctct gaatagattaatttaaagta gtcatgtaat 6240 gtttttttgg ttgcttacaa atgtcttttt attccaagcaggactataat atggatttag 6300 agctcgacga gtattataac aagacacttg ccacagagaataatactgct gccactcgga 6360 attctgattt cccagtctgg gatgactata aaagcagtgtagatgactta cagtattttc 6420 tgattgggct ctatacattt gtaagtcttc ttggctttatggggaatcta cttattttaa 6480 tggctctcat gaaaaagcgt aatcagaaga ctacggtaaacttcctcata ggcaatctgg 6540 ccttttctga tatcttggtt gtgctgtttt gctcacctttcacactgacg tctgtcttgc 6600 tggatcagtg gatgtttggc aaagtcatgt gccatattatgccttttctt caatgtgtgt 6660 cagttttggt ttcaacttta attttaatat caattgccattgtcaggtat catatgataa 6720 aacatcccat atctaataat ttaacagcaa accatggctactttctgata gctactgtct 6780 ggacactagg ttttgccatc tgttctcccc ttccagtgtttcacagtctt gtggaacttc 6840 aagaaacatt tggttcagca ttgctgagca gcaggtatttatgtgttgag tcatggccat 6900 ctgattcata cagaattgcc tttactatct ctttattgctagttcagtat attctgccct 6960 tagtttgtct tactgtaagt catacaagtg tctgcagaagtataagctgt ggattgtcca 7020 acaaagaaaa cagacttgaa gaaaatgaga tgatcaacttaactcttcat ccatccaaaa 7080 agagtgggcc tcaggtgaaa ctctctggca gccataaatggagttattca ttcatcaaaa 7140 aacacagaag aagatatagc aagaagacag catgtgtgttacctgctcca gaaagacctt 7200 ctcaagagaa ccactccaga atacttccag aaaactttggctctgtaaga agtcagctct 7260 cttcatccag taagttcata ccaggggtcc ccacttgctttgagataaaa cctgaagaaa 7320 attcagatgt tcatgaattg agagtaaaac gttctgttacaagaataaaa aagagatctc 7380 gaagtgtttt ctacagactg accatactga tattagtatttgctgttagt tggatgccac 7440 tacacctttt ccatgtggta actgatttta atgacaatcttatttcaaat aggcatttca 7500 agttggtgta ttgcatttgt catttgttgg gcatgatgtcctgttgtctt aatccaattc 7560 tatatgggtt tcttaataat gggattaaag ctgatttagtgtcccttata cactgtcttc 7620 atatgtaata attctcactg tttaccaagg aaagaacaaatgctggggtc atataaaata 7680 tatttatgat aactatttac atataataaa tagaaattttgttaacatgg aatttaattt 7740 atgtgaaaga gttctggatt caaatgtcag ttcataatatatggaagata attttatgtg 7800 ttatagtagg attaatttat ttagttgtgc agtcagtgtcaatccaatct gtaatttcac 7860 tttagaaggt tgtattacct tccacttcca tgttgtcttataaacaaatg aattgtattt 7920 tttgttgaaa gtaaaagtta tatctaacca actcagtacttttgtccaaa aatataataa 7980 gaaaaaattt ttctcgagga acttttaatt tcaaacttgaagaatatcta ccagctatct 8040 atatcatttc tactccatag gcttcttaat gtttagtttgtgaagtacag aaaaaattta 8100 atatgcctgg aaaatcacaa ctaaatgaca gatgtatgcccaaattatga ttataatctt 8160 caacattaac tacagttttg gaagtcctgt aggaaaatgctattgcctat tgagaattgg 8220 tcaaattgtc aatttaactc cactgtccta gtaatacacaagtaatttac caaataaaga 8280 attttaaatc ctttccagac tcattataca acattaaacactaccaataa aagttgtttt 8340 catatacatc aaaactattc taaaatgtga a 8371 2 445PRT Homo sapiens 2 Met Asp Leu Glu Leu Asp Glu Tyr Tyr Asn Lys Thr LeuAla Thr Glu 1 5 10 15 Asn Asn Thr Ala Ala Thr Arg Asn Ser Asp Phe ProVal Trp Asp Asp 20 25 30 Tyr Lys Ser Ser Val Asp Asp Leu Gln Tyr Phe LeuIle Gly Leu Tyr 35 40 45 Thr Phe Val Ser Leu Leu Gly Phe Met Gly Asn LeuLeu Ile Leu Met 50 55 60 Ala Leu Met Lys Lys Arg Asn Gln Lys Thr Thr ValAsn Phe Leu Ile 65 70 75 80 Gly Asn Leu Ala Phe Ser Asp Ile Leu Val ValLeu Phe Cys Ser Pro 85 90 95 Phe Thr Leu Thr Ser Val Leu Leu Asp Gln TrpMet Phe Gly Lys Val 100 105 110 Met Cys His Ile Met Pro Phe Leu Gln CysVal Ser Val Leu Val Ser 115 120 125 Thr Leu Ile Leu Ile Ser Ile Ala IleVal Arg Tyr His Met Ile Lys 130 135 140 His Pro Ile Ser Asn Asn Leu ThrAla Asn His Gly Tyr Phe Leu Ile 145 150 155 160 Ala Thr Val Trp Thr LeuGly Phe Ala Ile Cys Ser Pro Leu Pro Val 165 170 175 Phe His Ser Leu ValGlu Leu Gln Glu Thr Phe Gly Ser Ala Leu Leu 180 185 190 Ser Ser Arg TyrLeu Cys Val Glu Ser Trp Pro Ser Asp Ser Tyr Arg 195 200 205 Ile Ala PheThr Ile Ser Leu Leu Leu Val Gln Tyr Ile Leu Pro Leu 210 215 220 Val CysLeu Thr Val Ser His Thr Ser Val Cys Arg Ser Ile Ser Cys 225 230 235 240Gly Leu Ser Asn Lys Glu Asn Arg Leu Glu Glu Asn Glu Met Ile Asn 245 250255 Leu Thr Leu His Pro Ser Lys Lys Ser Gly Pro Gln Val Lys Leu Ser 260265 270 Gly Ser His Lys Trp Ser Tyr Ser Phe Ile Lys Lys His Arg Arg Arg275 280 285 Tyr Ser Lys Lys Thr Ala Cys Val Leu Pro Ala Pro Glu Arg ProSer 290 295 300 Gln Glu Asn His Ser Arg Ile Leu Pro Glu Asn Phe Gly SerVal Arg 305 310 315 320 Ser Gln Leu Ser Ser Ser Ser Lys Phe Ile Pro GlyVal Pro Thr Cys 325 330 335 Phe Glu Ile Lys Pro Glu Glu Asn Ser Asp ValHis Glu Leu Arg Val 340 345 350 Lys Arg Ser Val Thr Arg Ile Lys Lys ArgSer Arg Ser Val Phe Tyr 355 360 365 Arg Leu Thr Ile Leu Ile Leu Val PheAla Val Ser Trp Met Pro Leu 370 375 380 His Leu Phe His Val Val Thr AspPhe Asn Asp Asn Leu Ile Ser Asn 385 390 395 400 Arg His Phe Lys Leu ValTyr Cys Ile Cys His Leu Leu Gly Met Met 405 410 415 Ser Cys Cys Leu AsnPro Ile Leu Tyr Gly Phe Leu Asn Asn Gly Ile 420 425 430 Lys Ala Asp LeuVal Ser Leu Ile His Cys Leu His Met 435 440 445 3 2143 DNA Homo sapiens3 agctcgtcga cctgacctgc cacaaagtta gaagaaagga ttgattcaag aaagactata 60atatggattt agagctcgac gagtattata acaagacact tgccacagag aataatactg 120ctgccactcg gaattctgat ttcccagtct gggatgacta taaaagcagt gtagatgact 180tacagtattt tctgattggg gtctatacat ttgtaagtct tcttggcttt atggggaatc 240tacttatttt aatggctctc atgaaaaagc gtaatcagaa gactacggta aacttcctca 300taggcaatct ggccttttct gatatcttgg ttgtgctgtt ttgctgacct ttcacactga 360cgtctgtctt gctggatcag tggatgtttg gcaaagtcat gtgccatatt atgccttttc 420ttcaatgtgt gtcagttttg gtttcaactt taattttaat atcaattgcc attgtcaggt 480atcatatgat aaaacatccc atatctaata atttaacagc aaaccatggc tactttctga 540tagctactgt ctggacacta ggttttgcca tctgttctcc ccttccagtg tttcacagtc 600ttgtggaact tcaagaaaca tttggttcag cattgctgag cagcaggtat ttatgtgttg 660agtcatggcc atctgattca tagagaattg cctttactat ctctttattg ctagttcagt 720atattctgcc cttagtttgt cttactgtaa gtcatacaag tgtctgcaga agtataagct 780gtggattgtc caacaaagaa aacagacttg aagaaaatga gatgatcaac ttaactcttc 840atccatccaa aaagagtggg cctcaggtga aactctctgg cagccataaa tggagttatt 900cattcatcaa aaaacacaga agaagatata gcaagaagac agcatgtgtg ttacctgctc 960cagaaagacc ttctcaagag aaccactcca gaatacttcc agaaaacttt ggctctgtaa 1020gaagtcagct ctcttcatcc agtaagttca taccaggggt ccccacttgc tttgagataa 1080aacctgaaga aaattcagat gttcatgaat tgagagtaaa acgttctgtt acaagaataa 1140aaaagagatc tcgaagtgtt ttctacagac tgaccatact gatattagta tttgctgtta 1200gttggatgcc actacacctt ttccatgtgg taactgattt taatgacaat cttatttcaa 1260ataggcattt caagttggtg tattgcattt gtcatttgtt gggcatgatg tcctgttgtc 1320ttaatccaat tctatatggg tttcttaata atgggattaa agctgattta gtgtccctta 1380tacactgtct tcatatgtaa taattctcac tgtttaccaa ggaaagaaca aatgctgggg 1440tcatataaaa tatatttatg ataactattt acatataata aatagaaatt ttgttaacat 1500ggaatttaat ttatgtgaaa gagttctgga ttcaaatgtc agttcataat atatggaaga 1560taattttatg tgttatagta ggattaattt atttagttgt gcagtcagtg tcaatccaat 1620ctgtaatttc actttagaag gttgtattac cttccacttc catgttgtct tataaacaaa 1680tgaattgtat tttttgttga aagtaaaagt tatatctaac caactcagta cttttgtcca 1740aaaatataat aagaaaaaat ttttctcgag gaacttttaa tttcaaactt gaagaatatc 1800taccagctat ctatatcatt tctactccat aggcttctta atgtttagtt tgtgaagtac 1860agaaaaaatt taatatgcct ggaaaatcac aactaaatga cagatgtatg cccaaattat 1920gattataatc ttcaacatta actacagttt tggaagtcct gtaggaaaat gctattgcct 1980attgagaatt ggtcaaattg tcaatttaac tccactgtcc tagtaataca caagtaattt 2040accaaataaa gaattttaaa tcctttccag actcattata caacattaaa cactaccaat 2100aaaagttgtt ttcatataca tcaaaactat tctaaaatgt gaa 2143 4 445 PRT Homosapiens 4 Met Asp Leu Glu Leu Asp Asx Tyr Tyr Asn Lys Thr Leu Ala ThrGlu 1 5 10 15 Asn Asn Thr Ala Ala Thr Arg Asn Ser Asp Phe Pro Val TrpAsp Asp 20 25 30 Tyr Lys Ser Ser Val Asp Asp Leu Gln Tyr Phe Leu Ile GlyLeu Tyr 35 40 45 Thr Phe Val Ser Leu Leu Gly Phe Met Gly Asn Leu Leu IleLeu Met 50 55 60 Ala Leu Met Lys Lys Arg Asn Gln Lys Thr Thr Val Asn PheLeu Ile 65 70 75 80 Gly Asn Leu Ala Phe Ser Asp Ile Leu Val Val Leu PheCys Ser Pro 85 90 95 Phe Thr Leu Thr Ser Val Leu Leu Asp Gln Trp Met PheGly Lys Val 100 105 110 Met Cys His Ile Met Pro Phe Leu Gln Cys Val SerVal Leu Val Ser 115 120 125 Thr Leu Ile Leu Ile Ser Ile Ala Ile Val ArgTyr His Met Ile Lys 130 135 140 His Pro Ile Ser Asn Asn Leu Thr Ala AsnHis Gly Tyr Phe Leu Ile 145 150 155 160 Ala Thr Val Trp Thr Leu Gly PheAla Ile Cys Ser Pro Leu Pro Val 165 170 175 Phe His Ser Leu Val Glu LeuGln Glu Thr Phe Gly Ser Ala Leu Leu 180 185 190 Ser Ser Arg Tyr Leu CysVal Glu Ser Trp Pro Ser Asp Ser Tyr Arg 195 200 205 Ile Ala Phe Thr IleSer Leu Leu Leu Val Gln Tyr Ile Leu Pro Leu 210 215 220 Val Cys Leu ThrVal Ser His Thr Ser Val Cys Arg Ser Ile Ser Cys 225 230 235 240 Gly LeuSer Asn Lys Glu Asn Arg Leu Glu Glu Asn Glu Met Ile Asn 245 250 255 LeuThr Leu His Pro Ser Lys Lys Ser Gly Pro Gln Val Lys Leu Ser 260 265 270Gly Ser His Lys Trp Ser Tyr Ser Phe Ile Lys Lys His Arg Arg Arg 275 280285 Tyr Ser Lys Lys Thr Ala Cys Val Leu Pro Ala Pro Glu Arg Pro Ser 290295 300 Gln Glu Asn His Ser Arg Ile Leu Pro Glu Asn Phe Gly Ser Val Arg305 310 315 320 Ser Gln Leu Ser Ser Ser Ser Lys Phe Ile Pro Gly Val ProThr Cys 325 330 335 Phe Glu Ile Lys Pro Glu Glu Asn Ser Asp Val His GluLeu Arg Val 340 345 350 Lys Arg Ser Val Thr Arg Ile Lys Lys Arg Ser ArgSer Val Phe Tyr 355 360 365 Arg Leu Thr Ile Leu Ile Leu Val Phe Ala ValSer Trp Met Pro Leu 370 375 380 His Leu Phe His Val Val Thr Asp Phe AsnAsp Asn Leu Ile Ser Asn 385 390 395 400 Arg His Phe Lys Leu Val Tyr CysIle Cys His Leu Leu Gly Met Met 405 410 415 Ser Cys Cys Leu Asn Pro IleLeu Tyr Gly Phe Leu Asn Asn Gly Ile 420 425 430 Lys Ala Asp Leu Val SerLeu Ile His Cys Leu His Met 435 440 445 5 2286 DNA Rattus rattus 5gaattcggca cgaggggttt gcaaggtggc ttggaagtca actgccagta ggaaatagcc 60atccacacac ctgagttcca agggggaaga aagagattct tatctgattc tagtatggag 120tttaagcttg aggagcattt taacaagaca tttgtcacag agaacaatac agctgctgct 180cggaatgcag ccttccctgc ctgggaggac tacagaggca gcgtagacga tttacaatac 240tttctgattg ggctctatac attcgtaagt cttcttggct ttatgggcaa tctacttatt 300ttaatggctg ttatgaaaaa gcgcaatcag aagactacag tgaactttct cataggcaac 360ctggccttct ccgacatctt ggtcgtcctg ttttgctccc ctttcaccct gacctctgtc 420ttgttggatc agtggatgtt tggcaaaagc atgtgccata tcatgccgtt ccttcaatgt 480gtgtcagttc tggtttcaac tctgatttta atatcaattg ccattgtcag gtatcatatg 540ataaagcacc ctatttctaa caatttaacg gcaaaccatg gctacttcct gatagctact 600gtctggacac tgggctttgc catctgttct cccctcccag tgtttcacag tcttgtggaa 660cttaaggaga cctttgggct cagcactgct gagtagcaat atctctgtgt tgagtcatgg 720ccctctgatt catacagaat tgctttcaca atctctttat tgctagtgca gtatatcctg 780cctctagtat gtttaacggt aagtcatacc agcgtctgcc gaagcataag ctgtggattg 840tcccacaaag aaaacagact cgaagaaaat gagatgatca acttaaccct acagccatcc 900aaaaagagca ggaaccaggc aaaaaccccc agcactcaaa agtggagcta ctcattcatc 960agaaagcaca gaaggaggta cagcaagaag acggcctgtg tcttacccgc cccagcagga 1020ccttcccagg ggaagcacct agccgttcca gaaaatccag cctccgtccg tagccagctg 1080tcgccatcca gtaaggtcat tccaggggtc ccaatctgct ttgaggtgaa acctgaagaa 1140agctcagatg ctcatgagat gagagtcaag cgttccatca ctagaataaa aaagagatct 1200cgaagtgttt tctacagact gaccatactg atactcgtgt tcgccgttag ctggatgcca 1260ctccacgtct tccacgtggt gactgacttc aatgataact tgatttccaa taggcatttc 1320aagctggtat actgcatctg tcacttgtta ggcatgatgt cctgttgtct aaatccgatc 1380ctatatggtt tccttaataa tggtatcaaa gcagacttga gagcccttat ccactgccta 1440cacatgtcat gattctctct gtgcaccaaa gagagaagaa acgtggtaat tgacacataa 1500tttatacaga agtattctgg atctgaatgc cagttcgtaa tctacgtaag atcatcttca 1560tgttataata tggttaattc aatcagttgt gcagagtcaa tgtccatcta atacaatttc 1620atgtgttgaa gtagtttaca ttattttcca ttttatgtca ttggtaataa gttgagtgat 1680actctgtggt ttagtgtaaa atgtatgaag tgacaagttg tcccaaagag catttaacta 1740cagatttaag gaatttctat tatctgggta tcttcatttc tatttcacag gcttcttaac 1800atttttttgt aaaagtacaa aaatattcaa aagtcagaac tctattacag atgtatgcat 1860aaaagatgat tataattttg taggagaaag atctgctcct attagtgaag attggtaaaa 1920ttgtcagttt aacccgtctg tcctactact aatatttaat ttttcaaata tgaaaaggtt 1980tcagattttg tttagattta tatcacatta aacactgtca aataaaggct gtttttatat 2040gcatcgttga tgttccaaaa tgtgaagtct aaatggtgtc tgtatttcca attattaaat 2100aacttctaag atcattttta aaagtctgta gatggtatgg atagctagtt gtttgttaat 2160ataaagtaaa agtagatagc tgatttatgt tgtacctatg tcgtatgtat attaggagca 2220gtttcagccc cacagaacac tctatcgtgt tgtctcacta aagtgaaagc aaacgaaaaa 2280aaaaaa 2286 6 445 PRT Rattus rattus 6 Met Glu Phe Lys Leu Glu Glu HisPhe Asn Lys Thr Phe Val Thr Glu 1 5 10 15 Asn Asn Thr Ala Ala Ala ArgAsn Ala Ala Phe Pro Ala Trp Glu Asp 20 25 30 Tyr Arg Gly Ser Val Asp AspLeu Gln Tyr Phe Leu Ile Gly Leu Tyr 35 40 45 Thr Phe Val Ser Leu Leu GlyPhe Met Gly Asn Leu Leu Ile Leu Met 50 55 60 Ala Val Met Lys Lys Arg AsnGln Lys Thr Thr Val Asn Phe Leu Ile 65 70 75 80 Gly Asn Leu Ala Phe SerAsp Ile Leu Val Val Leu Phe Cys Ser Pro 85 90 95 Phe Thr Leu Thr Ser ValLeu Leu Asp Gln Trp Met Phe Gly Lys Ser 100 105 110 Met Cys His Ile MetPro Phe Leu Gln Cys Val Ser Val Leu Val Ser 115 120 125 Thr Leu Ile LeuIle Ser Ile Ala Ile Val Arg Tyr His Met Ile Lys 130 135 140 His Pro IleSer Asn Asn Leu Thr Ala Asn His Gly Tyr Phe Leu Ile 145 150 155 160 AlaThr Val Trp Thr Leu Gly Phe Ala Ile Cys Ser Pro Leu Pro Val 165 170 175Phe His Ser Leu Val Glu Leu Lys Glu Thr Phe Gly Ser Ala Leu Leu 180 185190 Ser Ser Lys Tyr Leu Cys Val Glu Ser Trp Pro Ser Asp Ser Tyr Arg 195200 205 Ile Ala Phe Thr Ile Ser Leu Leu Leu Val Gln Tyr Ile Leu Pro Leu210 215 220 Val Cys Leu Thr Val Ser His Thr Ser Val Cys Arg Ser Ile SerCys 225 230 235 240 Gly Leu Ser His Lys Glu Asn Arg Leu Glu Glu Asn GluMet Ile Asn 245 250 255 Leu Thr Leu Gln Pro Ser Lys Lys Ser Arg Asn GlnAla Lys Thr Pro 260 265 270 Ser Thr Gln Lys Trp Ser Tyr Ser Phe Ile ArgLys His Arg Arg Arg 275 280 285 Tyr Ser Lys Lys Thr Ala Cys Val Leu ProAla Pro Ala Gly Pro Ser 290 295 300 Gln Gly Lys His Leu Ala Val Pro GluAsn Pro Ala Ser Val Arg Ser 305 310 315 320 Gln Leu Ser Pro Ser Ser LysVal Ile Pro Gly Val Pro Ile Cys Phe 325 330 335 Glu Val Lys Pro Glu GluSer Ser Asp Ala His Glu Met Arg Val Lys 340 345 350 Arg Ser Ile Thr ArgIle Lys Lys Arg Ser Arg Ser Val Phe Tyr Arg 355 360 365 Leu Thr Ile LeuIle Leu Val Phe Ala Val Ser Trp Met Pro Leu His 370 375 380 Val Phe HisVal Val Thr Asp Phe Asn Asp Asn Leu Ile Ser Asn Arg 385 390 395 400 HisPhe Lys Leu Val Tyr Cys Ile Cys His Leu Leu Gly Met Met Ser 405 410 415Cys Cys Leu Asn Pro Ile Leu Tyr Gly Phe Leu Asn Asn Gly Ile Lys 420 425430 Ala Asp Leu Arg Ala Leu Ile His Cys Leu His Met Ser 435 440 445 71585 DNA Mus musculus 7 gttattgtca tagcgtgcta ttgttcttca agctgctaatggtcactgtc ttcttccaag 60 caggactcta gtatggaggt taaacttgaa gagcattttaacaagacatt tgtcacggag 120 aacaatactg ctgccagtca gaacacggcc tcccctgcctgggaggacta cagaggcaca 180 gagaacaata cttctgctgc tcggaacact ccgtttccagtctgggagga ctatagaggc 240 agcgtagacg acttacaata cttcctgatt gggctctatacatttgtaag tcttcttggt 300 tttatgggaa atctacttat cttaatggct gttatgaaaaagcgcaatca gaagactaca 360 gtgaactttc tcataggcaa cctggccttc tccgacattttggttgtcct gttttgctcc 420 cctttcaccc tgacctctgt cttgttggat cagtggatgttcggcaaagc catgtgccat 480 atcatgccat tccttcagtg tgtatcagtt ctggtttcaactctgatttt aatatcgatt 540 gccattgtca ggtatcatat gataaagcac cctatatctaacaatttaac agcaaaccat 600 ggctacttcc tgatagcatc tgtctggaca ctgggctttgccatctgttc tcccctccca 660 gtgtttcaca gccttgtgga acttaaggaa acctttggctcagcattgct aagcagcaag 720 tatttgtgtg ttgagtcatg gccctctgat tcatacagaattgctttcac aatctcttta 780 ttgttagttc agtatatcct gcctctagta tgtttaacagtaagtcatac tagtgtctgc 840 aggagtataa gctgtggatt gtcccacaaa gaaaacagactcgaagaaaa tgagatgatc 900 aacttaactc tacatccatc caaaaagagt cgggaccaggcaaaactccc cagcactcaa 960 aagtggagct actcattcat cagaaagcac cgaagaaggtacagcaagaa gacggcatgc 1020 gtgttacccg ccccagcagg accttcccag gagaagcacctaaccgttcc agaaaaccca 1080 ggctcggtcc gtagccagct gtcaccatcc agtaaggttattccaggggt cccgatctgc 1140 tttgaggtga aacctgaaga aagctcagat gctcaggagatgagagtcaa gcgttccctc 1200 acgagaataa agaagagatc tcgcagtgtt ttctacagactgactatatt gatattagtg 1260 ttcgctgtta gctggatgcc actccacgtc ttccacgtggtgaccgattt caatgataac 1320 ctgatttcca ataggcattt caagctggtg tactgcatctgtcacttgtt aggcatgatg 1380 tcctgttgtc ttaatccgat cttatatgga ttccttaataatggtatcaa agcagacttg 1440 agagccctta tccactgcct acacatgtca tgattctctctgtgcaccga ggagagaaga 1500 aatgtggaga ctgcccacaa tacatctgtg ctaattgatgcataatttac ataaacgtgt 1560 ctggatctga atgccagttt gtaat 1585 8 466 PRTMus musculus 8 Met Glu Val Lys Leu Glu Glu His Phe Asn Lys Thr Phe ValThr Glu 1 5 10 15 Asn Asn Thr Ala Ala Ser Gln Asn Thr Ala Ser Pro AlaTrp Glu Asp 20 25 30 Tyr Arg Gly Thr Glu Asn Asn Thr Ser Ala Ala Arg AsnThr Pro Phe 35 40 45 Pro Val Trp Glu Asp Tyr Arg Gly Ser Val Asp Asp LeuGln Tyr Phe 50 55 60 Leu Ile Gly Leu Tyr Thr Phe Val Ser Leu Leu Gly PheMet Gly Asn 65 70 75 80 Leu Leu Ile Leu Met Ala Val Met Lys Lys Arg AsnGln Lys Thr Thr 85 90 95 Val Asn Phe Leu Ile Gly Asn Leu Ala Phe Ser AspIle Leu Val Val 100 105 110 Leu Phe Cys Ser Pro Phe Thr Leu Thr Ser ValLeu Leu Asp Gln Trp 115 120 125 Met Phe Gly Lys Ala Met Cys His Ile MetPro Phe Leu Gln Cys Val 130 135 140 Ser Val Leu Val Ser Thr Leu Ile LeuIle Ser Ile Ala Ile Val Arg 145 150 155 160 Tyr His Met Ile Lys His ProIle Ser Asn Asn Leu Thr Ala Asn His 165 170 175 Gly Tyr Phe Leu Ile AlaSer Val Trp Thr Leu Gly Phe Ala Ile Cys 180 185 190 Ser Pro Leu Pro ValPhe His Ser Leu Val Glu Leu Lys Glu Thr Phe 195 200 205 Gly Ser Ala LeuLeu Ser Ser Lys Tyr Leu Cys Val Glu Ser Trp Pro 210 215 220 Ser Asp SerTyr Arg Ile Ala Phe Thr Ile Ser Leu Leu Leu Val Gln 225 230 235 240 TyrIle Leu Pro Leu Val Cys Leu Thr Val Ser His Thr Ser Val Cys 245 250 255Arg Ser Ile Ser Cys Gly Leu Ser His Lys Glu Asn Arg Leu Glu Glu 260 265270 Asn Glu Met Ile Asn Leu Thr Leu His Pro Ser Lys Lys Ser Arg Asp 275280 285 Gln Ala Lys Leu Pro Ser Thr Gln Lys Trp Ser Tyr Ser Phe Ile Arg290 295 300 Lys His Arg Arg Arg Tyr Ser Lys Lys Thr Ala Cys Val Leu ProAla 305 310 315 320 Pro Ala Gly Pro Ser Gln Glu Lys His Leu Thr Val ProGlu Asn Pro 325 330 335 Gly Ser Val Arg Ser Gln Leu Ser Pro Ser Ser LysVal Ile Pro Gly 340 345 350 Val Pro Ile Cys Phe Glu Val Lys Pro Glu GluSer Ser Asp Ala Gln 355 360 365 Glu Met Arg Val Lys Arg Ser Leu Thr ArgIle Lys Lys Arg Ser Arg 370 375 380 Ser Val Phe Tyr Arg Leu Thr Ile LeuIle Leu Val Phe Ala Val Ser 385 390 395 400 Trp Met Pro Leu His Val PheHis Val Val Thr Asp Phe Asn Asp Asn 405 410 415 Leu Ile Ser Asn Arg HisPhe Lys Leu Val Tyr Cys Ile Cys His Leu 420 425 430 Leu Gly Met Met SerCys Cys Leu Asn Pro Ile Leu Tyr Gly Phe Leu 435 440 445 Asn Asn Gly IleLys Ala Asp Leu Arg Ala Leu Ile His Cys Leu His 450 455 460 Met Ser 465

What is claimed is:
 1. An isolated DNA molecule encoding an NPY-Y5receptor consisting of about 445 amino acids, wherein said DNA moleculeencodes a mammalian NPY-Y5 receptor.
 2. An isolated DNA moleculeaccording to claim 1, wherein said DNA molecule encodes a human, mouseor rat NPY-Y5 receptor.
 3. An isolated DNA molecule according to claim2, wherein the DNA molecule encodes a human NPY-Y5 receptor.
 4. A methodfor detecting agonist or antagonist agents of NPY-Y5 receptor,comprising contacting a cell transformed with and expressing a DNAmolecule according to claim 1 with a test agent under conditionsenabling the activation of the NPY-Y5 receptor, and detecting anincrease or decrease in the NPY-Y5 receptor activity.
 5. An isolated DNAmolecule encoding an NPY-Y5 receptor, wherein the DNA molecule is atleast 95% identical to the nucleotide sequence shown: (i) at nucleotides6291 to 7625 of SEQ ID NO:1; (ii) at nucleotides 63 to 1397 of SEQ IDNO:3; (iii) at nucleotides 115 to 1449 of SEQ ID NO:5; or (iv) atnucleotides 73 to 1470 of SEQ ID NO:7.
 6. An isolated DNA moleculeencoding an NPY-Y5 receptor, wherein said DNA molecule comprises anucleotide sequence selected from the group consisting of nucleotides6291 to 7625 of SEQ ID NO:1, nucleotides 63 to 1397 of SEQ ID NO:3,nucleotides 115 to 1449 of SEQ ID NO:5, and nucleotides 73 to 1470 ofSEQ ID NO:7.
 7. The isolated DNA molecule encoding an NPY-Y5 receptoraccording to claim 6, wherein said DNA molecule comprises nucleotides6291 to 7625 of SEQ ID NO:1.
 8. The isolated DNA molecule encoding aNPY-Y5 receptor according to claim 6, wherein the DNA molecule comprisesnucleotides 63 to 1397 of SEQ ID NO:3.
 9. The isolated DNA moleculeencoding a NPY-Y5 receptor according to claim 6, wherein the DNAmolecule comprises nucleotides 115 to 1449 of SEQ ID NO:5.
 10. Theisolated DNA molecule encoding a NPY-Y5 receptor according to claim 6,wherein the DNA molecule comprises nucleotides 73 to 1470 of SEQ IDNO:7.
 11. A plasmid or expression vector comprising a DNA moleculeaccording to any one of claims 1 to 3 and 5 to
 10. 12. A host celltransformed with a DNA molecule according to any one of claims 1 to 3and 5 to
 10. 13. A host cell according to claim 12, wherein the cell isa mammalian or bacterial cell.
 14. A host cell according to claim 13,wherein the cell is a Chinese hamster ovary (CHO) cell or a humanembryonic kidney (HEK) 293 cell.
 15. A host cell according to claim 12,wherein the cell expresses NPY-Y5 receptor onto the cell's surface. 16.A host cell according to claim 12, wherein the cell is an insect Sf9cell.